Rifaximin prevents diclofenac-induced enteropathy in rats through antibacterial and anti-inflammatory activities

[Speaker] Matteo Fornai:1
[Co-author] Rocchina Colucci:2, Carolina Pellegrini:1, Erika Tirotta:1, Luca Antonioli:1, Cecilia Renzulli:3, Emilia Ghelardi:4, Elena Piccoli:4, Daniela Gentile:1, Laura Benvenuti:1, Gianfranco Natale:4, Federica Fulceri:1, Gloria Lopez-Castejon:5, Pablo Palazon-Riquelme:5, Corrado Blandizzi:1, Carmelo Scarpignato:6
1:Department of Clinical and Experimental Medicine, University of Pisa, Pisa, Italy, 2:Department of Pharmaceutical and Pharmacological Sciences, University of Padova, Padova, Italy, 3:Reasearch & Development Department, Alfasigma SpA, Bologna, Italy, 4:Department of Translational Research and New Technologies in Medicine and Surgery, University of Pisa, Pisa, Italy, 5:Manchester Collaborative Centre for Inflammation Research, University of Manchester, Manchester, UK, 6:Clinical Pharmacology & Digestive Pathophysiology Unit, Department of Clinical & Experimental Medicine, University of Parma, Parma, Italy

Background. Non-steroidal anti-inflammatory drugs can exert detrimental effects on the intestine, mainly through an involvement of enteric bacteria. This study examined the effects of rifaximin on small bowel mucosal integrity and inflammation as well as on gut microbiota in rats with diclofenac-induced enteropathy.
Methods. Enteropathy was induced in aged male rats by intragastric (i.g.) diclofenac (4 mg/kg BID, 14 days). Rifaximin was administered (50 mg/kg BID, i.g.) 1 hour before diclofenac. At the end of treatments, fecal calprotectin and blood hemoglobin levels were assayed (ELISA). Ileum was processed for the evaluation of: 1) histological damage; 2) tissue myeloperoxidase (MPO), tumor necrosis factor (TNF) and interleukin-1beta (IL-1beta) levels (ELISA); 3) quantitative analysis of bacterial strains (16S real-time PCR); 4) expression of toll-like receptors-2/-4 (TLR-2/-4), nuclear factor kB subunit p65 (p65), myeloid-differentiation primary response-gene 88 (MyD88), caspase-1 and occludin (Western blot). Moreover, the modulating effect of rifaximin on NLRP3 inflammasome signaling was tested in macrophagic THP-1 human cells.
Results. As compared with controls, rats treated with diclofenac displayed the occurrence of histological lesions in the ileum, along with decreased blood hemoglobin, and increased fecal calprotectin, tissue MPO, TNF and IL-1beta levels (Table). In control animals, the relative abundance of Proteobacteria, Firmicutes and Bacteroidetes was 4.8, 84.6 and 6.8%, respectively, while the prevalence of Lactobacilli was 68.5%. Diclofenac increased Proteobacteria and Bacteroidetes and decreased Firmicutes and Lactobacilli. The expression of TLR-2/-4, p65, MyD88 and activated caspase-1 in diclofenac-treated animals was higher, as compared with controls, while occludin expression was decreased. The co-administration of rifaximin reduced intestinal damage, inflammatory parameters (MPO, fecal calprotectin, TNF and IL-beta) and anemia induced by diclofenac (Table). In addition, rifaximin reduced the diclofenac-induced overexpression of TLR-2/-4, MyD88, p65 and the caspase-1 activation, while it counteracted the decrease in occludin expression. Rifaximin partly counteracted also the diclofenac-induced alterations of ileal microbiome. All parameters were not modified by rifaximin alone. In LPS-primed THP-1 cells stimulated with the selective NLRP3 activator nigericin, rifaximin concentration-dependently reduced IL-1beta production; this effect was associated with the inhibition of up-stream caspase-1 activation.
Conclusion. Rifaximin prevents diclofenac-induced enteropathy in rats through both antibacterial and anti-inflammatory activities.

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